METTL3调控结肠癌增殖、凋亡、浸润、转移和上皮间质化的研究

Study of METTL3 in proliferation,apoptosis,invasion,migration and epithelial-mesenchymal transition of colon cancer

目的探讨METTL3介导的m6A修饰在结肠癌中的作用。方法应用蛋白免疫印迹、免疫组化和实时荧光定量测定结肠癌细胞和组织中METTL3的表达水平。分别利用5-乙炔基-2′脱氧尿嘧啶核苷染色、TUNEL测定、Transwell实验和划痕实验评估METTL3对结肠癌细胞增殖、凋亡、浸润和转移的影响。应用蛋白免疫印迹测定E-钙黏蛋白、N-钙黏蛋白、TWIST和波形蛋白的表达水平,进而评价METTL3对结肠癌细胞上皮间质化的影响。结果与癌旁组织相比,METTL3在结肠癌组织表达水平显著升高(P<0.05)。下调METTL3的表达水平,不仅抑制结肠癌细胞的增殖(P<0.05)、浸润(P<0.05)和迁移(P<0.01),也诱导其凋亡(P<0.05)。另外,抑制METTL3的表达抑制结肠癌细胞上皮间质化:E-钙黏蛋白显著升高,而N-钙黏蛋白、波形蛋白和TWIST明显下降(P<0.05)。结论METTL3在结肠癌中可能发挥癌基因的作用。

Objective To investigate the role of methyltransferase-like 3(METTL3) mediated N6-methyladenosine(m6 A) in colon cancer. Methods Western blot, immunohistochemical(IHC) and quantitative real-time PCR(qRT-PCR) were used to detect METTL3 expression in colon cancer cell lines and patient tissues. The impact of METTL3 on cell proliferation, apoptosis, invasion and migration of colon cancer cells was examined by EdU staining, TUNEL staining, transwell assay and wound healing assay, respectively. The expression of E-Cadherin, N-Cadherin, Twist and Vimentin were measured by Western blot to evaluate the effect of METTL3 on EMT of colon cancer cells. Results The results showed higher METTL3 was expressed in colon cancer as compared with adjacent normal tissues(P<0.05). It was demonstrated that knocking down METTL3 inhibited the proliferation of colon cancer cells by cell proliferation experiment(P<0.05). TUNEL assay showed that downregulated METTL3 induced apoptosis of colon carcinoma cells(P<0.05). Transwell experiment and Wound healing assay indicated shRNA-METTL3 suppressed the invasion(P<0.05) and migration of SW620 cells(P<0.01). Moreover, Western blot analysis showed downregulated METTL3 upregulated the expression of E-cadherin and downregulated the level of N-cadherin, Vimentin and Twist(P<0.05). Conclusion METTL3, acting as an oncogene, might play a significant role in colon cancer tumorigenesis.